In the animal kingdom, as well as in some lower plants, i.e., nonvascular plants, fungi, yeast and bacteria, the primary reserve polysaccharide is glycogen. Glycogen is a polysaccharide containing linear molecules with .alpha.1, 4 glycosyl linkages and is branched via .alpha.1, 6 glycosyl linkages. Although from a linkage comparison, glycogen is analogous to starch, glycogen exhibits a different chain length and degree of polymerization. In bacteria, for example, the .alpha.1, 6 glycosyl linkages constitute only approximately 10% of the total linkages, indicating that the majority of the glycogen polymer resides as a linear glucose molecule.
In plants, i.e. vascular plants, reserve polysaccharides are stored in roots, tubers and seeds irk the form of starch. Starch, a complex polymer of glucose, consists of a mixture of linear chain and branched chain glucans known as amylose and amylopectin respectively. Starches isolated from different plants are found to have variable contents of amylose. Typically, amylose comprises from about 10-25% of plant starch, the remainder being the branched polymer amylopectin. Amylopectin contains low molecular weight chains and high molecular weight chains, with the low molecular weight chains ranging from 5-30 glucose units and the high molecular weight chains from 30-100 or more. The ratio of amylose/amylopectin and the distribution of low molecular weight to high molecular weight chains in the amylopectin fraction are known to affect the properties, such as thermal stabilization, retrogradation, and viscosity, and therefore utility of starch. The highest published low m.w./high m.w. chain ratios (on a weight basis) in amylopectin are 3.9/1 for waxy corn starch which has unique properties. Additionally, duwx, which has slightly more branch points than waxy has further unique properties.
In addition, starches from different plants or plant parts often have different properties. For example, potato starch has different properties than other starches, some of which may be due to the presence of phosphate groups. In some plant species, mutants have been identified which have altered contents of amylose and amylopectin. Mutations that affect the activity of starch-branching enzyme in peas, for example, result in seeds having less starch and a lower proportion of amylopectin. Also, mutations in the waxy locus of maize, which encodes a starch granule bound starch synthase, result in plants which produce amylopectin exclusively. Similarly, a potato mutant has been identified whose starch is amylose-free (Hovenkamp-Hermelink et al. Theor. Appl. Genet. (1987) 75:217-221). It has been found that varying the degree of starch branching can confer desirable physical properties; other changes in the characteristics of native starch could result in the production of polymers with new applications.
With the development of genetic engineering techniques, it is now possible to transfer genes from a variety of organism into the genome of a large number of different plant species. This process is preferable to plant breeding techniques whereby genes can only be transferred from one plant in a species to another plant in that same species or to a plant from a different, but closely related species. It would thus be desirable to develop plant varieties through genetic engineering, which have increased capacity for starch synthesis, altered amylose/amylopectin ratios, altered distribution of low to high molecular weight chains in the amylopectin fraction and also starches with novel molecular weight characteristics. In this manner, useful starches with a variety of viscosity or texture differences may be obtained.
To this end, nucleic acid sequences which encode glycogen biosynthetic enzymes are desirable for study and manipulation of the starch biosynthetic pathway. In particular, these enzymes may be expressed in plant cells using plant genetic engineering techniques and targeted to a plastid where starch synthesis occurs.